Autosomal-dominant polycystic kidney disease (ADPKD) is a common cause of end stage kidney disease. ADPKD is caused by mutations in one of two genes, PKD1 or PKD2, which are encoded by polycystin 1 (PC) and 2 respectively. Loss of both copies of PC1 or PC2 is associated with a decrease in of Ca2+ influx into mutant cells which is thought to mediate cyst formation and cyst enlargement by stimulating the enhanced growth of renal epithelia and the stimulation of apical chloride secretion via the cystic fibrosis transmembrane conductance regulator (CFTR). While the CFTR is directly regulated by cyclic AMP and is the predominant channel that secretes Cl- into the cyst lumen, we have evidence that a Ca2+-activated channel, KCa3.1, plays a critical role in CFTR-stimulated Cl- efflux in renal epithelia;by mediating the outflux of K+, KCa3.1 maintains the electrochemical driving force for Cl- secretion by setting the membrane potential at more negative values. KCa3.1 channels also play an important role in proliferation of a number of cells. Thus, inhibitors of KCa3.1 may serve a dual function to both inhibit the proliferation of renal epithelia and to inhibit Cl- secretion by the CFTR. The focus of this application is to study the role of KCa3.1 in the pathogenesis of PKD and to determine whether KCa3.1 is a viable drug target to slow disease progression. In Specific Aim (SA) 1 we will test: (i) whether inhibiting KCa3.1 by siRNA and by overexpression of a lipid phosphatase, myotubularin related protein 6 (MTMR6), inhibits Cl- secretion by MDCK cells;(ii) whether KCa3.1 is localized apically or basolaterally;(iii) whether direct activation of KCa3.1 by DCEBIO, stimulates Cl- secretion across an MDCK monolayer. In (B) we will determine whether genes known to affect KCa3.1 channel activity (MTMR6 and nucleoside diphosphate kinase B [NDPK-B]) function as modifiers to regulate Cl- secretion by the CFTR and cyst growth in vitro. In (C) we will extend the observations in SA1 A,B in MDCK cells to human and mouse renal tubule cells from wild type and PKD-/- cells and determine whether mutation in PKD1 or PKD2 affects KCa3.1 regulation, function, or activity. In SA2, we will determine the relevance of KCa3.1 to cyst formation in mouse models of PKD1 and PKD2. We will determine whether treatment of mice with TRAM-34, a specific inhibitor of KCa3.1, blocks cyst formation and progression to renal failure in mice models for PKD1 and PKD2. Autosomal-dominant polycystic kidney disease affects is a common cause of end stage kidney disease. Over time, these cysts become more numerous and larger in size and replace normal kidney tissue leading to loss of renal function. In this proposal, we are studying a potassium channel that we have evidence is important for the movement of salt and water into the cyst lumen which is thought to be one of primary mechanism whereby cysts enlarge over time. In addition, this channel may also play an important role in proliferation or growth of these cells, which is also important for cyst formation. The excitement in studying this channel is that drugs that inhibit this channel (KCa3.1) already exists and are in human trials without any major side effects. Thus, if inhibiting this channel shows promise in mouse models of ADPKD, we can rapidly move into the clinic to assess treatment in patients.